PROJECT SUMMARY It is estimated that in the United States alone there will be nearly 70 million people over the age of 65 by the year 2030. Yet, there remains a critical unmet need to understand and identify novel therapeutic strategies to address age-related impairments in arousal, cognitive function, and sleep architecture. Agents such as acetylcholinesterase inhibitors (AChEIs) that enhance overall cholinergic transmission or activate muscarinic acetylcholine receptors (mAChRs) have been developed to ameliorate the loss of cognitive function in normal aging and Alzheimer?s disease (AD) populations. Accumulating evidence indicates that enhancement of central cholinergic neurotransmission through activation of the M1 mAChR subtype represents an exciting alternative approach for the treatment of impairments in cognition, arousal and sleep observed in normal aging and AD. M1 is co-localized with the NR1a subunit of the N-methyl-D-aspartate subtype of glutamate receptor (NMDAR). Selective activation of M1 potentiates prefrontal cortical (PFC) and hippocampal NMDAR currents and enhances both PFC- and hippocampal-mediated associative learning and memory functions. Recently we successfully optimized a novel series of isoindolinone M1 positive allosteric modulators (PAMs), represented by VU0453595, as an alternative strategy for the selective activation of M1. The discovery and characterization of VU0453595 represents an unprecedented opportunity to characterize the impact of selective M1 potentiation on cortical ACh neurotransmission and cognition, arousal and sleep dysfunction associated with normal aging in preclinical species. In Aim 1, we will determine effects of VU0453595 alone and in combination with the AChEI donepezil on cortical acetylcholine and glutamate neurotransmission using in vivo microdialysis techniques in young and aged male and female mice. Next, we will examine effects of VU0453595 alone and in combination with donepezil to ameliorate age-related impairments in cognition and sleep architecture in aged male and female mice (Aim 2). Finally, in Aim 3 we will investigate whether selective activation of M1 by VU0453595 ameliorates age-related impairments in cognition and sleep architecture in male and female aged NHPs. Together, these translational studies across species will extend our understanding of age-related cognitive dysfunction, a critical risk factor for later onset dementia and AD, and the potential role of M1 modulation for restoration of age-related impairments in cognition and sleep architecture.